Purification and characterization of astrocyte-secreted apolipoprotein E and J-containing lipoproteins from wild-type and human apoE transgenic mice
Purification and characterization of astrocyte-secreted apolipoprotein E and J-containing lipoproteins from wild-type and human apoE transgenic mice
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DOI：
10.1016/s0197-0186(01)00049-3
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发表时间：
2001-11-01
影响因子：
4.2
通讯作者：
Holtzman, DM
中科院分区：
文献类型：
--
作者：
DeMattos, RB;Brendza, RP;Holtzman, DM
载脂蛋白E（apoE）的ε 4等位基因是阿尔茨海默病（AD）的遗传危险因素。为了更好地了解apoE和其他可能的载脂蛋白在中枢神经系统（CNS）中产生的影响AD发病机制的分子机制，我们已经纯化并表征了两种最丰富的载脂蛋白在CNS中产生，apoE和apoJ。我们从小鼠星形胶质细胞的原代培养物中纯化了apoE和apoJ，所述星形胶质细胞来源于在小鼠apoE不存在的情况下表达人apoE同种型的转基因小鼠。利用抗体亲和柱，我们能够纯化人载脂蛋白E3和载脂蛋白E4，以及小鼠载脂蛋白J。星形胶质细胞分泌的人载脂蛋白E存在于高密度样脂蛋白的三个主要尺寸范围从8到15 nm的直径。小鼠apoJ是直径在10和17 nm之间的颗粒，峰值大小范围类似于11 nm。ApoE和apoJ在不同的脂蛋白中。利用快速冷冻，深蚀刻电子显微镜显示，apoE颗粒盘，而apoJ颗粒更小，更不规则的外观。载脂蛋白E颗粒的脂质组成与含有载脂蛋白J的颗粒有很大不同。载脂蛋白E颗粒含有类似质量的载脂蛋白E和脂质。其中胆固醇和磷脂在每颗粒的质量上大约相等。ApoJ-颗粒相对缺乏脂质（三部分蛋白质。一部分脂质），磷脂比胆固醇丰富得多。通过电喷雾电离质谱分析的磷脂组合物的详细表征揭示乙醇胺甘油磷脂是apoE和apoJ颗粒中存在的最丰富的磷脂。apoE 3和apoE 4转基因小鼠的脑脊液分析显示，人类和小鼠apoE的颗粒大小与星形胶质细胞分泌的颗粒相同。CNS衍生脂蛋白的生理制剂的进一步使用可以允许详细了解这些分子在正常脑和疾病如AD中的作用。(C)，2001爱思唯尔科技有限公司版权所有。
The epsilon4 allele of apolipoprotein E (apoE) is a genetic risk factor for Alzheimer's disease (AD). In order to gain a better understanding of the molecular mechanisms by which apoE and possibly other apolipoproteins produced in the central nervous system (CNS) influence AD pathogenesis, we have purified and characterized the two most abundant apolipoproteins produced in the CNS, apoE and apoJ. We purified apoE and apoJ from primary cultures of mouse astrocytes, which were derived from transgenic mice expressing human apoE isoforms in the absence of mouse apoE. Utilizing antibody affinity columns, we were able to purify both human apoE3 and apoE4, as well as mouse apoJ-containing lipoproteins. Astrocyte-secreted human apoE was present in high density-like lipoproteins of three predominant sizes ranging from 8 to 15 nm in diameter. Mouse apoJ was in particles between 10 and 17 nm in diameter with a peak size range of similar to 11 nm. ApoE and apoJ were in distinct lipoproteins. Utilization of quick-freeze, deep-etch electron microscopy revealed the apoE particles were discs while the apoJ particles were smaller and more irregular in appearance. The lipid composition of apoE particles was very different from those containing apoJ. ApoE-particles contained a similar mass of apoE and lipid. with cholesterol and phospholipid being about equal in mass per particle. ApoJ-particles were relatively lipid poor (three parts protein. one part lipid), with phospholipids being much more abundant than cholesterol. Detailed characterization of phospholipid composition by electrospray ionization mass spectrometry analysis revealed ethanolamine glycerophospholipids to be the most abundant phospholipid present in both apoE and apoJ particles. Analysis of cerebrospinal fluid from apoE3 and apoE4 transgenic mice revealed that human and mouse apoE were in particles the same size as those secreted by astrocytes. Further use of physiological preparations of CNS-derived lipoproteins may allow for a detailed understanding of the role of these molecules in the normal brain and in diseases such as AD. (C), 2001 Elsevier Science Ltd. All rights reserved.